Last data update: 24 January 2024 16:39 CET
Plasmid name: pDG2i-mRipk3-Flag puro (LMBP 10386)
| New search | Print data sheet |
| Price category: | Cat. 1 (cf. price list) |
| Status: | GeneCorner core plasmid |
| GeneCorner sequence: |
p10386.gb
(View with Genome Compiler) p10386.txt p10386.pdf |
| Sequence analysis results Genecorner: |
- |
| Cloned DNA: |
Mouse receptor-interacting serine-threonine kinase 3 cDNA (Ripk3, Rip3, GeneID 56532) Reverse tetracycline-responsive transcriptional activator (rtTA); variant 3 (rtTA3) FLAG epitope tag; C-terminal |
| Promoter: | Human cytomegalovirus immediate early promoter (CMV-IE); minimal promoter with tet responsive element (CMVTRE) Rous sarcoma virus/human immunodeficiency virus hybrid long terminal repeat (RSV/HIV-1 5' LTR) Simian virus 40 early promoter (SV40 early) Escherichia coli lac operon promoter Phage T3 promoter Phage T7 gene 10 promoter (T7g10) |
| Ribosome binding site: |
Internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV) polyprotein gene |
| Terminator: | Rous sarcoma virus/human immunodeficiency virus hybrid long terminal repeat (RSV/HIV-1 3' LTR); modified HIV-1 3' LTR with deleted U3 region (ΔU3/3' LTR) Simian virus 40 polyadenylation signal (SV40 polyA) |
| Selection marker: | Ampicillin (amp) Puromycin (puro) |
| Replicon: | Simian virus 40 bidirectional origin (SV40) Escherichia coli plasmid pMB1 origin Phage f1 origin |
| Host range: | Escherichia coli Mammalian cells; SV40 permissive cells |
| Parental clone: | pDG2i-Flag puro; pDONOR 207-mRIP3K |
| Further information: | The plasmid was constructed by cloning the mouse Rip3 coding sequence into the pDG2i-Flag puro vector via Gateway LR recombination with pDONOR 207-mRIP3K. This is a lentiviral Gateway expression vector, with the tet-inducible minimal human CMV-IE promotor and rtTA3 on the same plasmid. The U3 region of the HIV-1 3' LTR is deleted (ΔU3 or dU3) and facilitates self-inactivation of the HIV-1 5' LTR after transduction to enhance the biosafety of the vector. The element also contains a polyadenylation signal for transcription termination and polyadenylation of mRNA in transduced cells. The vector contains the psi packaging signal, allowing viral packaging, and the rev response element (RRE) from HIV-1, permitting Rev-dependent nuclear export of unspliced viral mRNA. Because the risk for recombination after each subcultivation is high, this plasmid is only available under the format of isolated plasmid DNA. Plasmid integrity should be checked via PvuII restriction digest after each subcultivation. The nucleotide sequence of the parental pDG2i-Flag puro vector corresponds with the EMBL Nucleotide Sequence Database accession number LT009444.1. Other names of the plasmid are pDG2'-'Flag rtTA3 puro + mRIP3K_FL cl2 and pDG2-Flag-rtTA3-puro-mRIPK3. |
| EMBL Accession number: | LT009444.1, view at EMBL, GenBank, DDBJ |
| Latest sequence update: | 26/07/2017 |
| Authenticity test: | Restriction enzyme pattern analysed at GeneCorner: HindIII, NdeI, NdeI/SpeI, NdeI/XhoI, PvuII, SpeI/XhoI. |
| Class: | Recombinant plasmid |
| Type: | Plasmid |
| History of deposit: | The plasmid was deposited by Dr T. L. Aaes(1)(2) and Prof. Dr P. Vandenabeele(1)(2). (1) Center for Inflammation Research, VIB, Ghent, Belgium (2) Department of Biomedical Molecular Biology, Ghent University, Ghent, Belgium |
| Plasmid reference: | Aaes et al., Cell Rep. 15 (2016), 274-287 [PMID: 27050509] [DOI: 10.1016/j.celrep.2016.03.037] |
| Related plasmid reference: | De Groote et al., BioTechniques 60 (2016), 252-259 [PMID: 27177818] [DOI: 10.2144/000114417] |
| Restricted distribution: | - BCCM MTA - RECIPIENT agrees to refer to the 'Plasmid reference' in the first publication making use of the MATERIAL. |
| Distributed as: | plasmid DNA |
| Host for distribution: | Escherichia coli K12 DH5α |
| Host reference: | Focus 8 (1986), 9 |
| Related host reference: | Woodcock et al., Nucleic Acids Res. 17 (1989), 3469-3478 [PMID: 2657660] Rodriguez-Quinones et al., Focus 15 (1993), 110-112 Hanahan, J. Mol. Biol. 166 (1983), 557-580 [PMID: 6345791] [DOI: 10.1016/s0022-2836(83)80284-8] Hanahan, in 'DNA Cloning: A Practical Approach Volume I', IRL Press, Oxford (1985), 109-135 [ISSN/ISBN: 0947946187] Grant et al., Proc. Natl. Acad. Sci. U.S.A. 87 (1990), 4645-4649 [PMID: 2162051] |
| Cultivation medium: | LB-Lennox + ampicillin (100 μg/ml) |
| Cultivation temperature: | 28°C |
| Biosafety level: | L1 in E. coli; L2 in mammalian cells |
| Other culture collection numbers: | - |
Refer in your Materials and Methods: |
pDG2i-mRipk3-Flag puro (LMBP 10386) is available at BCCM/GeneCorner. The plasmid was deposited by Dr T. L. Aaes and Prof. Dr P. Vandenabeele and was published in Aaes et al., 2016. |
Note: Up-to-date, validated data are enclosed with the biological material. Nevertheless, these data are a snapshot at a given moment; further updates are always possible.